This project will develop, optimize and implement technology to lineally mark and/or genetically alter individual germ cells in C. elegans. The germ line in this worm shares many features in common with stem cell systems in all animals, including humans. For example, it is maintained by virtue of interaction with a cellular microenvironment or niche that produces ligands to activate a highly conserved Notch receptor on the germ cells. It was the first stem cell system for which a niche was defined at the cell and molecular levels. This stem cell system has since emerged as a powerful system to understand the dynamics of stem cell expansion, quiescence and plasticity in response to genetic, environmental and physiological alterations. The worm has a short (3-day) lifecycle, is transparent, short-lived, and highly amenable to environmental and genetic manipulation. Despite these advantages, understanding critical features of the germline stem cell system and its responses to environmental and genetic perturbations is currently limited by our relatively poor understanding of cellular mechanisms of renewal and how they are altered under experimental manipulation. To bridge this gap, we need to follow individual stem cells and their cellular progeny over time. The technology proposed here will bridge this gap. Taking advantage of recent progress in germline transgene expression in C. elegans, existing technologies from other organisms will be brought to the worm. In addition to lineage marking, the technology will make possible the manipulation of gene activity in single cells or subsets of cells. Uniquely for this model system, the technology will enable monitoring of germline stem cells in individuals over their entire lifetime and in conditions of whole-animal physiological perturbations. This advance will benefit the greater stem cell biology and germline fields as additional questions will become accessible in this premier model system.